Parathyroid hormone-related protein (PTHrP) and the Type 1 parathyroid hormone/parathryoid hormone related protein receptor (PTH1R) are required for the formation of mammary glands in mouse and human embryos. From work funded by the first cycle of this grant, we now know that PTHrP acts as a signal from the epithelial cells of the mammary bud to the surrounding mesenchyme that promotes the differentiation of a mammary-specific condensed mesenchyme, maintains the mammary fate of the epithelial cells, instructs the overlying keratinocytes to form the nipple and supports outgrowth of the bud into the initial duct system. In this cycle, will plan to study the molecular mechanisms by which PTHrP signaling accomplishes these developmental events. We have also found that PTHrP is made by terminal end-buds in the growing epithelial ducts of the mammary gland during puberty, while the PTH1R is expressed in the surrounding stromal cells. Our data suggest that PTHrP inhibits the effects of estrogen on ductal growth. In this cycle, we will explore the mechanisms by which PTHrP modulates the effects of estrogen on mammary stromal cells during puberty. In order to pursue these goals, we propose three specific aims. The first will examine the interactions between PTHrP and BMP signaling in the embryonic mammary bud. The second will use gene array technology to define the mesenchymal factors that are responsible for promoting ductal outgrowth in response to PTHrP signaling. The third will seek to establish whether endogenous PTHrP acts to restrain ductal growth in response to estrogen, and will explore the potential interactions between PTHrP signaling and estrogen-induced epidermal growth factor receptor activation in mammary stromal cells. The processes involved in breast cancer progression and metastasis are in many ways similar to those involved n embryonic and pubertal mammary development. We hope that our studies will contribute to a better understanding of both normal mammary development and the pathogenesis of breast cancer.